Background: Sentinel lymph node (SLN) location and biopsy was designed to minimize side effects of axillary dissection with equivalent outcomes. Aim: To evaluate the feasibility of periareolar injection of contrast agent SonoVueTM followed by ultrasound (US) for identification and localization of SLN in breast cancer patients with clinically negative node. Methods: From July 2017 through January 2018, 130 women were enrolled in the study. SonoVueTM was injected periareolarly and followed by US to detect enhanced sentinel lymphatic channels (SLCs) and SLNs 1 minute later after massage. The patients were randomly divided into two groups to locate the first enhanced SLN: 1) US-guided marker placing; 2) US-guided nano-carbon (N) injection into SLN. Compare the number of SLNs detect by CEUS with blue dye (B) or N mapping, and the coincidence rate of the first SLN located by CEUS with those traced by the B and N. Lymph nodes that were dark, blue, with marker or clinically positive were considered sentinel nodes and to be biopsied. Results: 121 of 130 patients with breast cancer patients injected with ultrasound contrast agents had detected a total of 254 enhanced SLNs (range 1-5, 2.1 ± 1.05) compared with total of 342 SLNs (range 1-5, 2.83 ± 1.10) mapping with B or N. 42 of 45 first SLNs located with marker matched with first SLNs stained with N (42/45, 93.33%); 70 of 76 first SLNs using N injecting directly guided by CEUS matched with first SLNs stained with B (70/76, 92.1%). In another 9 cases without enhanced SLN, 4 of them didn't stained with N or B, and 5 were stained. The sensitivity of SLNs detection by CEUS was 96.03% (121/126), and the accuracy of locating the first SLN was 92.56% (112/121). Conclusion: CEUS has good accuracy in the detection and localization of SLNs in patients with breast cancer.